During the luteal phase of the primate menstrual cycle, the corpus luteum produces steroid hormones including progesterone. Serum progesterone levels are low in early luteal phase, peak at midluteal phase, and fall to very low levels by late luteal phase, indicating that progesterone production by the corpus luteum is regulated during the luteal phase. The final step in progesterone synthesis is catalyzed by the enzyme 3b-hydroxysteroid dehydrogenase (3b-HSD), and expression of this enzyme in the monkey corpus luteum has been confirmed using immunocytochemistry. To determine if changes in luteal 3b-HSD mRNA levels correlate with changes in serum progesterone, a riboprobe antisense to macaque 3b-HSD mRNA was developed and used for Northern analysis of macaque total RNA. 3b-HSD mRNA was detected in steroidogenic tissues, such as the ovary and adrenal, but not in non-steroidogenic tissues, such as liver, heart, and spleen. The presence of 3b-HSD mRNA in monkey luteal tissue was confirmed. A linear relationship between the amount of total RNA and 3b-HSD signal suggests that Northern blotting can be used for quantitative analysis of 3b-HSD mRNA. Initial studies using monkey luteal RNA from tissues collected at various stages of the luteal phase suggest that 3b-HSD mRNA levels parallel changing serum progesterone levels. Evidence from several species suggests that progesterone acts at the corpus luteum to regulate its own production. We treated monkeys with the 3b-HSD inhibitor trilostane to reduce serum progesterone to very low levels and determined that progesterone depletion may reduce luteal 3b-HSD mRNA levels. These data suggest that progesterone regulates its own production through modulation of steroidogenic enzyme gene expression in the primate corpus luteum.